Facultad de Ciencias Quimicas, Universidad Autonoma de Nuevo Leon, Avenida Universidad s/n, Ciudad Universitaria, 66455, San Nicolas de los Garza, NL, Mexico.
Departamento de Patologia Clinica, Hospital Universitario Dr. Jose Eleuterio Gonzalez, Universidad Autonoma de Nuevo Leon, 64460, Monterrey, NL, Mexico.
Appl Microbiol Biotechnol. 2024 Dec;108(1):126. doi: 10.1007/s00253-023-12887-5. Epub 2024 Jan 13.
An alarming global public health and economic peril has been the emergence of antibiotic resistance resulting from clinically relevant bacteria pathogens, including Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumonia, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter species constantly exhibiting intrinsic and extrinsic resistance mechanisms against last-resort antibiotics like gentamycin, ciprofloxacin, tetracycline, colistin, and standard ampicillin prescription in clinical practices. The discovery and applications of antimicrobial peptides (AMPs) with antibacterial properties have been considered and proven as alternative antimicrobial agents to antibiotics. In this study, we have designed, produced, and purified a recombinant novel multifunctional hybrid antimicrobial peptide LL-37_Renalexin for the first time via the application of newly designed flexible GS peptide linker coupled with the use of our previously characterized small metal-binding proteins SmbP and CusF3H+ as carrier proteins that allow for an enhanced bacterial expression, using BL21(DE3) and SHuffle T7(DE3) Escherichia coli strains, and purification of the hybrid peptide via immobilized metal affinity chromatography. The purified tag-free LL-37_Renalexin hybrid peptide exhibited above 85% reduction in bacteria colony-forming units and broad-spectrum antimicrobial effects against Staphylococcus aureus, Escherichia coli, Methicillin-resistant Staphylococcus aureus (MRSA), and Klebsiella pneumoniae bacteria clinical isolates at a lower minimum inhibition concentration level (10-33 μM) as compared to its counterpart single-AMPs LL-37 and Renalexin (50-100 μM). KEY POINTS: • The hybrid antimicrobial peptide LL-37_Renalexin has been designed using a GS linker. • The peptide was expressed with the carrier proteins SmbP and CusF3H+. • The hybrid peptide shows antibacterial potency against clinical bacterial isolates.
一种令人震惊的全球公共卫生和经济危害是，临床相关细菌病原体（包括屎肠球菌、金黄色葡萄球菌、肺炎克雷伯菌、鲍曼不动杆菌、铜绿假单胞菌和肠杆菌属）对抗生素的固有和获得性耐药性不断出现，这些细菌对临床实践中最后使用的抗生素如庆大霉素、环丙沙星、四环素、黏菌素和标准氨苄西林产生耐药性。具有抗菌特性的抗菌肽（AMPs）的发现和应用已被认为是抗生素的替代抗菌药物。在这项研究中，我们首次通过应用新设计的柔性 GS 肽接头，将小金属结合蛋白 SmbP 和 CusF3H+ 与我们之前表征过的载体蛋白结合，设计、生产和纯化了一种新型多功能杂合抗菌肽 LL-37_Renalexin，该方法可增强细菌表达，使用 BL21(DE3) 和 SHuffle T7(DE3) 大肠杆菌菌株，并通过固定化金属亲和层析对杂合肽进行纯化。纯化的无标签 LL-37_Renalexin 杂合肽对金黄色葡萄球菌、大肠杆菌、耐甲氧西林金黄色葡萄球菌（MRSA）和肺炎克雷伯菌临床分离株的抑菌效果提高了 85%以上，最低抑菌浓度（MIC）水平低于其对应的单 AMPs LL-37 和 Renalexin（50-100 μM）。 关键点： • 设计了使用 GS 接头的杂合抗菌肽 LL-37_Renalexin。 • 该肽与载体蛋白 SmbP 和 CusF3H+ 一起表达。 • 杂合肽对临床分离的细菌具有抗菌效力。
